NanoDam identifies Homeobrain (ARX) and Scarecrow (NKX2.1) as conserved temporal factors in the Drosophila central brain and visual system

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Spatial and temporal patterns of neurogenesis in the central nervous system of Drosophila melanogaster

Neurogenesis in the ventral CNS of Drosophila was studied using staining with toluidine blue and birth dating of cells monitored by incorporation of bromodeoxyuridine into DNA. The ventral CNS of the larva contains sets of neuronal stem cells (neuroblasts) which are thought to be persistent embryonic neuroblasts. Each thoracic neuromere has at least 47 of these stem cells whereas most abdominal neuromeres possess only 6. They occur in stereotyped locations so that the same neuroblast can be followed from animal to animal. The thoracic neuroblasts begin enlarging at 18-26 hr of larval life, DNA synthesis commences by 31-36 hr, and the first mitoses occur shortly thereafter. Mitotic activity continues through the remainder of larval life with the neuroblasts showing a minimum cell cycle time of less than 55 min during the late third larval instar. By 12 hr after pupariation each neuroblast has produced approximately 100 progeny which are collected with it into a discrete packet. The progeny accumulate in an immature, arrested state and only finish their differentiation into mature neurons with the onset of metamorphosis. Most of the abdominal neuroblasts differ from their thoracic counterparts in their minimum cell cycle time (less than 2 hr) and the duration of proliferation (from about 50 to 90 hr of larval life). Neurons produced during the larval stage account for more than 90% of the cells found in the ventral CNS of the adult.     

Mutations in the Drosophila neuroglian cell adhesion molecule affect motor neuron pathfinding and peripheral nervous system patterning

We have identified and characterized three embryonic lethal mutations that alter or abolish expression of Drosophila Neuroglian and have used these mutations to analyze Neuroglian function during development. Neuroglian is a member of the immunoglobulin superfamily. It is expressed by a variety of cell types during embryonic development, including expression on motoneurons and the muscle cells that they innervate. Examination of the nervous systems of neuroglian mutant embryos reveals that motoneurons have altered pathfinding trajectories. Additionally, the sensory cell bodies of the peripheral nervous system display altered morphology and patterning. Using a temperature-sensitive mutation, the phenocritical period for Neuroglian function was determined to occur during late embryogenesis, an interval which coincides with the period during which neuromuscular connections and the peripheral nervous system pattern are established. © 1997 John Wiley & Sons, Inc. J Neurobiol 32: 325–340, 1997     

Genetic control of the developmental program of L-glycerol-3-phosphate dehydrogenase isozymes in Drosophila melanogaster: Identification of a cis-acting temporal element affecting GPDH-3 expression

The complete developmental program of glycerol-3-phosphate dehydrogenase in wild type Drosophila is described with respect to activity, isozyme expression, and GPDH-specific CRM. Variants of this developmental program have been isolated from natural populations which affect the rate of accumulation of only the GPDH-3 isozyme in both the larval and adult stages of development. This activity variation segregates as a single gene which is tightly linked to the structural element on Chromosome II, exhibits cis-control, and is tissue specific in expression. This gene meets all the criteria for temporal regulatory genes.     

Characterization of dendritic spines in the Drosophila central nervous system

Dendritic spines are a characteristic feature of a number of neurons in the vertebrate nervous system and have been implicated in processes that include learning and memory. In spite of this, there has been no comprehensive analysis of the presence of spines in a classical genetic system, such as Drosophila, so far. Here, we demonstrate that a subset of processes along the dendrites of visual system interneurons in the adult fly central nervous system, called LPTCs, closely resemble vertebrate spines, based on a number of criteria. First, the morphology, size, and density of these processes are very similar to those of vertebrate spines. Second, they are enriched in actin and devoid of tubulin. Third, they are sites of synaptic connections based on confocal and electron microscopy. Importantly, they represent a preferential site of localization of an acetylcholine receptor subunit, suggesting that they are sites of excitatory synaptic input. Finally, their number is modulated by the level of the small GTPase dRac1. Our results provide a basis to dissect the genetics of dendritic spine formation and maintenance and the functional role of spines. © 2009 Wiley Periodicals, Inc. Develop Neurobiol, 2009     

Mechanisms of early neurogenesis in Drosophila melanogaster

The neuroectoderm of insects contains an initially indifferent population of cells which during later development will give rise to the progenitor cells of the neural and epidermal lineages. Experimental evidence indicates that cellular interactions determine which cells will adopt each one of these fates. Transplantation experiments suggest that a signal with neuralising character is required to stabilize the primary neural fate in 25% of all the neuroectodermal cells, which will develop as neuroblasts, and that an epidermalising signal contributes to suppress the neural fate in the remaining 75% of the cells, allowing in this way their development as epidermal progenitor cells. The invoked cell interactions are assumed to be mediated by the products of several genes forming a complex, not yet well understood network of interrelationships. Elements of this network are the proteins encoded by Delta and Notch, which appear to convey the regulatory signals between the cells; the proteins encoded by the achaete-scute gene complex, which regulate neural development; and the proteins encoded by the Enhancer of split gene complex, which give neuroectodermal cells access to epidermal development. © 1993 John Wiley & Sons, Inc.     

Spatial regulation of segment polarity gene expression in the anterior terminal region of the Drosophila blastoderm embryo

The effects of mutations in five anterior gap genes (hkb, tll, otd, ems and btd) on the spatial expression of the segment polarity genes, wg and hh, were analyzed at the late blastoderm stage and during subsequent development. Both wg and hh are normally expressed at blastoderm stage in two broad domains anterior to the segmental stripes of the trunk region. At the blastoderm stage, each gap gene acts specifically to regulate the expression of either wg or hh in the anterior cephalic region: hkb, otd and btd regulate the anterior blastoderm expression of wg, while tll and ems regulate hh blastoderm expression. Additionally, btd is required for the first segmental stripe (mandibular segment) of both hh and wg at blastoderm stages. The subsequent segmentation of the cephalic segments (preantennal, antennal and intercalary) appears to be dependent on the overlap of the wg and hh cephalic domains as defined by these gap genes at the blastoderm stage. None of these five known gap genes are required for the activation of the labral segment domains of hh and wg, which are presumably either activated directly by maternal pathways or by an unidentified gap gene.     

Stage-specific regulation of actin genes in Drosophila wing cells

Extreme and rapid changes in the synthesis of messenger RNAs and proteins accompany differentiation in wing tissues of Drosophila. Of the six actin genes, at least three are expressed in wing cells, some during the most extreme changes in cell shape. However, different messages of the set appear, decay, and reappear on a regulated temporal program. These results show that actin expression is stage-specific in a single cell type.     

模式动物果蝇的基因调控前沿技术

转基因调控技术在生命医学研究中扮演了重要的角色,是探究个体发育和致病机制的必备工具。常用的转基因调控技术包括基因突变、基因干扰和基因转录激活等。果蝇由于具有基因的保守性、遗传工具的多样性以及不受伦理限制等优势,成为生命科学研究中经典模式动物之一,并由此开发出了多种时间和组织特异性的基因调控工具。本文主要介绍了目前在果蝇中常用的基因调控技术,包括CRISPR/Cas9介导的基因突变系统、基于miRNA的新一代转基因干扰系统以及基于CRISPR/dCas9的转录激活(flySAM)系统,希望通过对这几种系统设计原理、操作过程、相关的技术工具及相关资源品系的介绍,提升人们对这些前沿的果蝇遗传学基因表达调控技术及构建的相关果蝇资源品系重要性认识,进而推动生命医学研究发展。     

Cell lineage analysis of the Drosophila peripheral nervous system

The peripheral nervous system (PNS) of Drosophila provides a very well-characterized model system for studying the genes involved in basic processes of neurogenesis. Because of its simplicity and stereotyped pattern, each cell of the PNS can be individually identified and the phenotypic consequences of mutations can be studied in detail. Thus, some of the genetic mechanisms leading to the formation of type I sensory organs, the external, bristle-type sensory organs (es), and the internal, stretch-receptive chordotonal organs (ch) have been elucidated. Each sensory organ seems to be generated by a stereotyped pattern of cell division of individual ectodermal precursor cells. Recent advances in cell lineage analysis of the PNS have provided a detailed picture of almost all the lineages in the PNS, including those giving rise to the type II sensory neurons, also known as multiple dendritic (md) neurons. This knowledge will be instrumental in the precise characterization of the phenotypes associated with mutations in known and new genes and their interactions which determine cell fate decisions during neurogenesis. Here, we describe and compare three recently developed methods by which cell lineages have been assessed: single cell transplantation, bromodeoxyuridine (BrdU) incorporation studies, and the flp/FRT recombinase system from yeast. In the light of a more complete knowledge of the PNS lineages, we will discuss the effects of known mutations that alter neuronal cell fates. © 1996 Wiley-Liss, Inc.     

Glial cells phagocytose neuronal debris during the metamorphosis of the central nervous system in Drosophila melanogaster

 Using electron microscopy we demonstrate that degenerating neurons and cellular debris resulting from neuronal reorganization are phagocytosed by glial cells in the brain and nerve cord of the fruitfly Drosophila melanogaster during the first few hours following pupariation. At this stage several classes of glial cells appear to be engaged in intense phagocytosis. In the cell body rind, neuronal cell bodies are engulfed and phagocytosed by the same glial cells that enwrap healthy neurons in this region. In the neuropil, cellular debris in tracts and synaptic centres resulting from metamorphic re-differentiation of larval neurons is phagocytosed by neuropil-associated glial cells. Phagocytic glial cells are hypertrophied, produce large amounts of lysosome-like bodies and contain a large number of mitochondria, condensed chromatin bodies, membranes and other remains from neuronal degeneration in phagosomes. Received: 23 January 1996 / Accepted in revised form: 21 May 1996     

Regulatory interactions during early neurogenesis in Drosophila

During neurogenesis in Drosophila, ectodermal cells are endowed with the capacity to become neuronal precursors. Following their selection, these cells initiate neuronal lineage development and differentiation. The processes of neuronal precursor specification and neuronal lineage development require the activities of several groups of genes functioning in a complex, hierarchical regulatory network. Whereas the proneural genes promote neurogenic potential, neurogenic genes restrict the acquisition of this identity to a subset of ectodermal cells. Following their selection, these cells express the pan neural neuronal precursor genes and a set of neuronal lineage identity genes. While lineage identity genes allow the various lineages to acquire specific identities, neuronal precursor genes presumably regulate functional and developmental characteristics common to all neuronal precursor cells. © 1996 Wiley-Liss, Inc.     

Drosophila highwire gene modulates acute ethanol sensitivity in the nervous system

Animals exhibit behavioral differences in their sensitivity to ethanol, a trait that is at least in part due to genetic predispositions. This study has implicated a large neuronal protein involving Highwire, a Drosophila E3 ubiquitin ligase (Hiw, a homolog of Pam, a protein associated with Myc found in humans) in acute sensitivity to ethanol sedation. Flies lacking Hiw were hypersensitive to the sedating effect of ethanol whereas those overexpressing Hiw showed decreased sensitivity to ethanol. Furthermore, RNAi functional knockdown of Hiw in adult neurons or ellipsoid body neurons showed increased sensitivity to ethanol sedation. None of these manipulations of the hiw gene caused changes in the rate of ethanol absorption and/or metabolism. These results suggest a previously unknown role for this highly conserved gene in regulating the behavioral responses to an addictive drug.     

Domain specific genetic mosaic system in the Drosophila eye

Genetic mosaic approach is commonly used in the Drosophila eye by completely abolishing or misexpressing a gene within a subset of cells to unravel its role during development. Classical genetic mosaic approach involves random clone generation in all developing fields. Consequently, a large sample size needs to be screened to generate and analyze clones in specific domains of the developing eye. To address domain specific functions of genes during axial patterning, we have developed a system for generating mosaic clones by combining Gal4/UAS and flippase (FLP)/FRT system which will allow generation of loss‐of‐function as well as gain‐of‐function clones on the dorsal and ventral eye margins. We used the bifid‐Gal4 driver to drive expression of UAS‐FLP. This reagent can have multiple applications in (i) studying spatio‐temporal function of a gene during dorso‐ventral (DV) axis specification in the eye, (ii) analyzing genetic epistasis of genes involved in DV patterning, and (iii) conducting genome wide screens in a domain specific manner. genesis 51:68–74, 2013. © 2012 Wiley Periodicals, Inc.     

Drosophila larval neuromuscular junction's responses to reduction of cAMP in the nervous system

We investigated the effects of chronically lowered cyclic adenosine monophosphate (cAMP) on the morphology and physiology of the Drosophila larval neuromuscular junction, using two fly lines in which cAMP was significantly lower than normal in the nervous system: (a) transgenic flies in which the dunce (dnc) gene product was overexpressed in the nervous system, and (b) flies mutant for the rutabaga gene (rut¹) which have reduced adenylyl cyclase activity. In comparison with controls, larvae with reduced cAMP exhibited a smaller number of synaptic varicosities. This effect was more pronounced in transgenic larvae, in which the reduction of neural cAMP was more pronounced. Synaptic transmission was also reduced in both cases, as evidenced by smaller excitatory junctional potentials (EJPs). Synaptic currents recorded from individual synaptic varicosities of the neuromuscular junction indicated almost normal transmitter release properties in transgenic larvae and a modest impairment in rut¹ larvae. Thus, reduction in EJP amplitude in transgenic larvae is primarily due to reduced innervation, while in rut¹ larvae it is attributable to the combined effects of reduced innervation and a mild impairment of transmitter release. We conclude that the major effect of chronically lowered cAMP is reduction of innervation rather than impairment of transmitter release properties. © 1999 John Wiley & Sons, Inc. J Neurobiol 40: 1–13, 1999